Synapses are specialized sites of cell-cell contact that mediate communication between neurons in the nervous system. Much remains to be discovered about the molecular mechanisms that underlie formation of these critical structures in the mammalian central nervous system. Previously, we developed a novel, forward genetic, RNA-interference (RNAi)-based approach to identifying new molecules that regulate synapse formation. Currently, we are in the process of applying this technology to understanding the cell biology of drug addiction. Thus far, a number of kinases have been implicated in regulating synapse formation or function, lending support to the hypothesis that protein kinases have critical functions at the synapse. Further, activation of protein kinase signaling has been hypothesized to underlie changes in neuronal structure and synapses in response to drug exposure. Therefore, further investigation into the role of protein kinases in synapse formation is warranted. To this end, we propose to take a genome-wide approach to identify the full complement of protein kinases that are expressed at the time that synapses are forming in cultured mammalian neurons. Next, we will utilize our RNAi-based screening approach to ask which kinases are required for the formation of functional glutamatergic and/or GABAergic synapses. PUBLIC HEALTH RELEVANCE: A current hypothesis to explain the persistent features of drug addiction, including drug cravings and relapse, posits that changes in synaptic structure and neuronal connectivity underlie these features of the disease. Further, the function of protein kinases has been implicated in these changes in synaptic structure. Thus, a genome-wide approach to understanding the role of protein kinases in synapse formation and function as outlined in this proposal has the potential to yield important insights into the underlying causes of a subset of features of drug addiction.